The objectives of this proposal will seek to develop and characterize a model system for the targeting of toxins to specific cells which have the potential for inducing tumors in mammals. Derivatives of the octapeptide toxin amanitin i.e. Alpha-amanitin coupled via diazotization with benzoic acid, aminobenzoyl glycine, and aminobenzoylglycylglycine; o-methyl-aldo-Alpha-amanitin coupled via reductive aldo condensation with Gamma-aminobutyric acid (Gamma-ABA) and glycylglycine (GG); Beta-amanitin via aminde bonds with Gamma-ABA and GG; 6'dehydroxyamanullinic acid vis aminde bonds with Gamma-ABA and GG, will be synthesized to permit covalent conjugation via active esters to free amino groups on monoclonal antibodies directed against cell surface antigens. Conjugates will be prepared with each derivative and monoclonal antiThy-1.2 IgG. One or more effective derivatives will be conjugated to antiLyt-1.12 IgG, antiLyt-2.2 IgG, andantiThy-1.2 Fab, (Fab')2, and IgM. Amanitin derivatives will be coupled to adjunct carriers; i.e. polymers of L-lysine, for increased loading of antibody conjugates. The conjugates will be characterized with respect to the nature of the amanitin-protein linkage, the number of amanitin moieties bound, and the inhibitory activity of the coupled amanitin toward calf thymus RNA polymerase II and cell specific cytotoxicity in vitro. Labeled conjugates (3H-amanitin, 125I-protein) will be utilized to determine the mechanism of endocytosis and processing of select conjugates under conditions which lead to specific cell inhibition. conjugates with antiThy-1.2 Fab, (Fab')2, IgG, and IgM will be examined for their ability to block the proliferation of S 49.1 (Thy 1.2+) and S 49 (thy-) cells inoculated in syngeneic Balb/c, and in nude mice. The effect of injection of these conjugates on the survival of inoculated mice will be determined. The excretion of 3H and 125I by mice injected with conjugates containing a given 3H derivative of amanitin and 125I antiThy-1.2 (Fab, (Fab')2, IgG, and/or IgM) will be determined to evaluate residence times and clearance. Labeled conjugates will be quantified in localized tumors and tissues to determine the extent of targeting.